Septohippocampal system plays an important role in regulating fear and anxiety behaviors. In this study, the effects of histamine injected into the dorsal hippocampus and opioidergic agents into medial septum on the anxiety-like behaviors in rats were analyzed, using the Elevated Plus-Maze (EPM) test. Injection of 1 and 5 μg/rat histamine into dorsal hippocampus had no effect on anxiety-like behavior, while injection of 10 μg/rat histamine increased the percentage of open arm time (%OAT) and open arm entry (%OAE), which indicated the anxiolytic effects of histamine. Microinjection of morphine, μ-opioid receptor agonist, into the medial septum (1μg/rat) increased the (%OAT) and (%OAE). Doses of 0.25, 0.5 μg/rat morphine had no effect on anxiety. Co-administration of histamine ineffective dose (1μg/rat) to the dorsal hippocampus and ineffective dose of morphine (0.25µg/rat) to the medial septum increased the (%OAT) and (%OAE). Subsequently, injection of different doses of naloxone (1, 2, 4 µg/rat), as an opioid receptor antagonist, into the medial septum in the presence and absence of an effective dose of histamine (10 µg/rat) in the dorsal hippocampus, was studied. Injection of naloxone (4 µg/rat) into medial septum decreased the (%OAT) and (%OAE), but did not alter the locomotor activity, which indicated the anxiogenic effects of naloxone. Simultaneous injection of histamine (10 µg/rat) into dorsal hippocampus with doses of naloxone (1, 2, 4 µg/rat) into the medial septum, indicate anxiolytic effects and increased %OAT and %OAE in Elevated Plus Maze, although when the dose of naloxone was 4μg/rat, this effect was less observed. The results indicate that hippocampus histaminergic system interact with medial septum opioidergic system and the interaction of these systems modulates anxiety behavior.